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A sensitive ELISA made with recombinant antibodies for cynomolgus IL-8 in serum
Leting Yang1, Xia Han2, Xiao Zhu2
1State Key Laboratory of Discovery and Utilization of Functional Components in Traditional Chinese Medicine & School of Pharmaceutical Sciences, Guizhou Medical University, Guiyang, 561113, China; Zhongshan Institute for Drug Discovery, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Zhongshan, 528400, China.
Accurate measurement of interleukin-8 (IL-8) in non-human primates is crucial for research into inflammation and tumors; however, sensitive enzyme-linked immunosorbent assays (ELISAs) for non-clinical studies remain limited. We aimed to develop and validate a sensitive method for IL-8 quantification in cynomolgus monkey serum. Using two recombinant IL-8-specific antibodies and a surrogate matrix, a sandwich-type ELISA was developed, involving sequential incubations with samples, biotinylated antibody, and horseradish peroxidase-labeled Streptavidin (SA-HRP). The assay was validated against regulatory guidelines, achieving a lower limit of quantification of 4.69 pg/mL as well as good accuracy and precision. The assay was neither affected by serum matrix nor hemolysis, nor did it cross-react with other cytokines. Furthermore, the assay showed dilution linearity, no hook effect, and good recovery and parallelism. Finally, IL-8 in monkey serum exhibited sufficient stability for analysis. After validation, the assay was applied to 19 healthy cynomolgus monkey serum samples, and the results correlated well with those from a commercial assay. In conclusion, the developed ELISA is a sensitive, accurate, and reliable method for IL-8 quantification in cynomolgus monkey serum, suitable for non-clinical studies in cynomolgus monkeys.
Accurate measurement of interleukin-8 (IL-8) in non-human primates is crucial for research into inflammation and tumors; however, sensitive enzyme-linked immunosorbent assays (ELISAs) for non-clinical studies remain limited. We aimed to develop and validate a sensitive method for IL-8 quantification in cynomolgus monkey serum. Using two recombinant IL-8-specific antibodies and a surrogate matrix, a sandwich-type ELISA was developed, involving sequential incubations with samples, biotinylated antibody, and horseradish peroxidase-labeled Streptavidin (SA-HRP). The assay was validated against regulatory guidelines, achieving a lower limit of quantification of 4.69 pg/mL as well as good accuracy and precision. The assay was neither affected by serum matrix nor hemolysis, nor did it cross-react with other cytokines. Furthermore, the assay showed dilution linearity, no hook effect, and good recovery and parallelism. Finally, IL-8 in monkey serum exhibited sufficient stability for analysis. After validation, the assay was applied to 19 healthy cynomolgus monkey serum samples, and the results correlated well with those from a commercial assay. In conclusion, the developed ELISA is a sensitive, accurate, and reliable method for IL-8 quantification in cynomolgus monkey serum, suitable for non-clinical studies in cynomolgus monkeys.
